Mycobacterium avium infections are still a major clinical concern in HIV infected individuals. Like other pathogenic mycobacteria, M. avium reside in macrophages, where they can avoid being killed by the immune system. In the first aim, we propose to continue our studies on the identification of potential M. avium subunit vaccines. We have previously shown that the FAP-1 protein is immunogenic in mice and immune responses to it are detected in both infected mice and humans. We propose to examine the ability of he FAP-A protein and identified epitopes to provide protective immunity in M. avium infections. These immunogens will be administered as purified proteins or expressed in attenuated Salmonella. In aims 2 and 3 we propose to utilize a novel strategy to identify genes which are induced upon entry of the M. avium into a macrophage. This strategy is based upon the hypothesis that a mycobacterium upon entry into a macrophage senses its environment and turns on a cascade of genes, which permit it to survive inside the macrophage and avoid recognition by the immune system. To identify these candidate genes, we propose to use a two-component regulatory system as our probe. These extensively studied regulatory systems are utilized by essentially all bacteria to sense and respond to their environments. A two-component regulatory system has been identified in M. tuberculosis, which is stimulated upon entry into a macrophage. We have cloned the equivalent two-component system in M. avium, named marA/B. We will examine the entire mycobacteria genome for sequences and genes which are regulated by marA, using a genetic screen. Our initial studies will examine the M. tuberculosis genome, but what we will use the M. avium genome as soon as it becomes available. From these studies, we will identify candidate genes which are involved in the survival of M. avium inside macrophages. We will then directly examine the role of marA/B system and the candidate genes in pathogenesis. We will generate mycobacterial strains deficient in the gene of interest and then test their growth inside macrophages and their in vivo pathogenesis. These identified genes will also be excellent candidates for novel drug targets. Overall, these studies should help to identify the mechanism(s) by which a M. avium can reside in macrophage and avoid being killed by the immune system.